The long term objective is to understand the eukaryotic nucleus, using the yeast, Saccharomyces cerevisiae, as a model system. The goal is to identify and characterize genes affecting the structure and function of chromosomes. Current effort is focused on understanding the mechanism of transcriptional silencing in yeast. This serves as a model for understanding heterochromatin in metazoans. We will study the role of the N-terminal BAH domain present in the silencing protein Sir3, as well as in the replication protein Orel. Random mutations are being introduced into these domains in order to study the function of particular subdomains. In particular, the very N-terminus, identical in Sir3 and Orel, will be studied structurally by crystallography and functionally by genetic analysis. We also want to understand why a particular surface of the nucleosome, near H3 Lys79, is so important for silencing by determining which Sir protein binds to this region of the nucleosome. Finally, we want to understand the role of sumoylation of Lys247 of Sir3 and its interaction with the ring finger proteins, Ris1 and Hex3/Slx5. Progress in these areas will shed light on chromosome structure and function in the eukaryotic nucleus, not only in yeast but also in mammalian cells.